1. Field of the Invention
This invention relates to spectroscopy, in particular, devices and methods for spectrographic analyses that use filters to select wavelengths of electromagnetic radiation for measurement.
2. Description of Related Art
Spectroscopy can be characterized as the study of relationships between absorption and/or emission of electromagnetic radiation by certain substances as a function of the wavelength of the radiation. Absorption spectroscopy is in widespread use for the detection and identification of substances because a substance absorbs electromagnetic radiation better at certain wavelengths than at others. When a substance is exposed to a poorly absorbed wavelength of electromagnetic radiation, much of that radiation is reflected or transmitted back into the surrounding medium. A photodetector nearby can detect the radiation, and the amount of radiation can be quantified. In contrast, when a substance is exposed to an efficiently absorbed wavelength, little of that radiation is reflected into the surrounding medium, and consequently, the amount of radiation detected is less than for a poorly absorbed wavelength. Measurements are typically made over a range of wavelengths, and can include very short wavelengths (e.g., gamma- rays or x-rays) to very long wavelengths (e.g., radio frequency radiation). The relationship between radiation intensity and wavelength is herein termed a xe2x80x9cspectrum.xe2x80x9d As used here, the term xe2x80x9cspectrumxe2x80x9d includes, but is not limited to absorption, fluorescence, Raman, emission, or any other form or type of electromagnetic radiation. For many analytical applications, wavelengths in ultraviolet, visible and/or infrared ranges are especially useful.
Individual substances either absorb or emit characteristic wavelengths of electromagnetic radiation. Each substance thus has a characteristic spectrum, which can be used to identify and/or quantify the amount of a particular substance. Many volumes in the spectroscopic literature are devoted to the presentation of data regarding spectra of individual substances.
However, existing methods and apparatus have several drawbacks. Most spectroscopic apparatus rely upon varying the wavelength of emitted radiation from a radiation source by means of a dispersion device such as a prism or a diffraction grating. A dispersion device decomposes electromagnetic radiation of heterogeneous wavelengths into spatially resolved beams of fairly monochromatic radiation. The dispersion is achieved as follows: An electromagnetic radiation is collimated in a beam to allow the beam to fall onto a prism or grating under appropriate angle of incidence. Radiation of various wavelengths present in the beam interferes with such a dispersion device in a wavelength-dependent manner. This produces a plurality of fairly monochromatic beams radiated under various, wavelength-dependent angles. Each beam is collected onto the surface of a photosensitive device (such as a photo-multiplying tube, also called PMT, or photo-diode, or photo-sensitive film). The intensity of monochromatic light in such a beam is analyzed as the function of spatial position of the beam. The position is directly related to the wavelength in the beam. This way of spectra acquisition is broadly employed in various spectrophotometers and spectrographs. A major drawback of this approach is a high cost for such instrumentation, which is to a large extend due to a need for precise alignment of optical elements.
A source of electromagnetic radiation (e.g., a light source) produces a beam of radiation that enters a dispersion device. By way of example, a prism separates the different wavelengths at different angles depending on the index of refraction of each wavelength as it is transmitted through the prism. In the case of visible light, the result can be a xe2x80x9crainbow.xe2x80x9d To expose an analyte sample to a particular wavelength, the prism is adjusted so that the angle of refraction of the radiation directs a relatively narrow range of wavelengths to the sample for spectroscopic measurement. To obtain a spectrum, the wavelength is varied by rotating the prism to direct other wavelengths to the sample. Similar methods can be applied to diffraction gratings. These processes are relatively slow, in that the rate of change of wavelength of illuminating radiation must be sufficiently slow to permit accurate measurement of absorption at each wavelength.
The length of time required to obtain a spectrum over a desired range of wavelengths depends upon the range desired, the discrimination between wavelengths, and upon the number of samples to be analyzed. For analyses of multiple samples, traditional spectroscopic methods can be impractically long. Moreover, prisms and diffraction gratings must be aligned carefully and misalignment can result in errors that may be difficult to detect.
To overcome these and other disadvantages of traditional spectroscopic devices and methods, certain embodiments of this invention use a plurality of narrow-band pass filters to select wavelengths of electromagnetic radiation for analysis. Each filter can be associated with an individual detector, for example, a charge coupled device (xe2x80x9cCCDxe2x80x9d), forming a xe2x80x9cfilter/detector unitxe2x80x9d. Radiation emitted by a sample can penetrate through a filter and can be detected and/or quantified and can be displayed on an output device and/or stored in electronic form on a computer. The filter can absorb radiation of other wavelengths, preventing those wavelengths from being detected. Additional filters having desired transmittance at other, selected wavelengths can be used simultaneously to detect absorption at those desired wavelengths.
Multiple filter/detector units can be placed in a one- or two-dimensional arrangement relative to each other, permitting the simultaneous measurement of absorbed radiation at a number of different wavelengths from a single sample of the substance to be analyzed. Outputs from each detector can be displayed along, for example, a vertical axis of a two-dimensional plot, and the band-pass wavelength of the filter can be displayed along a horizontal axis, for example, similar to a conventional spectrogram. Thus, a spectrum can be obtained over a desired range of wavelengths. Addressable arrays of samples can be analyzed in an automated fashion. A series of samples can be applied to a substrate, each sample having a unique identifier, either position on the array, or by way of a unique chemical marker. Systems for spectrographic analysis can include servo-controlled probes that can acquire spectrographic information from each of a plurality of samples so arrayed.
It can be readily appreciated that similar strategies can be employed for emission, fluorescence, Raman, and any other kind of spectra, and other types of plots (e.g., three-dimensional displays) can be readily prepared.
In certain embodiments, filters can be chosen to permit passage of a relatively narrow wavelength band of radiation. Such embodiments can be useful in situations in which a desired spectrographic feature is narrow.
In certain other embodiments, filters can be chosen to permit passage of a relatively wide wavelength band of radiation. Such embodiments can be useful in situations in which desired spectrographic features are broad, or in which the desired information has sufficiently high intensity and is not masked by signals at other wavelengths within the band detected.
In yet other embodiments, a portion of a spectrum can be obtained using filter/detector units having wavelength bands that are sufficiently near each other to provide substantially complete coverage throughout a desired wavelength range. In other embodiments, it can be desirable to select only certain portions of a spectrum for analysis.
In additional embodiments of this invention, filter/detector units can include waveguides, including light pipes to transmit radiation from a sample to a remote detector.
Many configurations of sample, sample substrate, waveguides, focusing lenses and detectors are possible. In certain embodiments, a plurality of samples can be prepared on a substrate in an array, and samples can be xe2x80x9creadxe2x80x9d sequentially.
Certain embodiments employ lenses or other means to focus radiation emitted by a sample onto a waveguide for transmission to a detector. Focusing can increase the intensity of the signal detected and/or can decrease the amount of radiation arising from other samples in an array (xe2x80x9cparasite radiationxe2x80x9d) which can confound the analysis of certain spectrographic features.
Spectrographic information from small samples or a portion of a sample can be obtained using the above strategy along with microscopes. Resolution of microscopic detection of spectra can depend upon the wavelengths of interest, with features in low wavelength portions of the electromagnetic spectrum (e.g., violet/ultraviolet) permitting finer detail than for features having longer wavelengths (e.g., infrared).
In other embodiments, the filters can be miniaturized and arranged in a one- or a two-dimensional array to permit the simultaneous measurement of absorption at different wavelength bands of a relatively small sample.
In yet other embodiments of this invention, arrays of miniaturized filter/detector units can be formed as a probe and can be positioned sequentially over different samples. Such embodiments can be especially desired for spectrographic analysis of multiple samples on a substrate.
In yet further embodiments, a plurality of arrays of miniaturized filter/detector units can be used simultaneously to obtain spectrographic analyses of a multiplicity of samples simultaneously.
In certain other embodiments, the filters can be of fixed band-pass, or alternatively, in other embodiments, can be made xe2x80x9ctunablexe2x80x9d using electric field-sensitive liquid crystal materials and/or any other materials possessing the desired, similar optical and/or electrical properties.
The apparatus and methods of this invention can avoid many of the problems facing conventional spectrophotometric methods and apparatus. In situations in which the different filters have fixed wavelength band ranges, the problems of optical alignment can be reduced. Because such filters can be made reproducibly, wavelength drift can be minimized. Moreover, the lack of a requirement for sophisticated moving parts can permit manufacture of relatively inexpensive, yet accurate spectrographic devices.
The use of multiple filter/detector units can permit the simultaneous measurement of a desired spectrum or portion thereof, which can substantially reduce the length of time required for spectrographic analyses. By providing accurate rapid analyses, the devices and methods of this invention can permit study of volatile and/or fragile analytes. By way of example, an analyte that is easily vaporized can be detected sufficiently rapidly to permit acquisition of a broad range of wavelengths simultaneously. In contrast, prior art dispersion based methods can suffer from artifacts in the spectrum due to loss of sample during the analysis. Specifically, later-measured wavelengths can have artificially low signal intensity due to loss of the analyte, and the true relationship between peak intensities can be misrepresented. Similarly, for analytes that are labile, i.e., that are fragile and can degrade easily, the devices and methods of this invention can provide improved spectra. As with volatile analytes, prior art dispersion based methods and devices can result in later-measured wavelengths being under represented relative to earlier-measured wavelengths. Moreover, using the devices and methods of this invention, spectra can be obtained under a variety of different ambient conditions including reduced temperature and/or chemical milieu. Thus, conditions can be selected that can reduce artifacts and result in more accurate, reproducible spectrographic analyses.
Devices and methods of this invention can be used for analyte detection, identification of substances for materials science applications, and astrophysical studies of radiation emitted by remote objects. For example, gamma-radiation and x-radiation can provide important information concerning stars, galaxies quasars, neutron stars and other astrophysical phenomena. Infrared and/or radio frequency detectors can be useful for studying features opaque to visible radiation, including surface features of planets having atmospheres.